The Medical Research Council Mitochondrial Biology Unit, University of Cambridge, Keith Peters Building, Cambridge Biomedical Campus, Hills Road, Cambridge CB2 0XY, UK.
Sci Adv. 2023 Aug 2;9(31):eadi1359. doi: 10.1126/sciadv.adi1359.
Respiratory complex I, a key enzyme in mammalian metabolism, captures the energy released by reduction of ubiquinone by NADH to drive protons across the inner mitochondrial membrane, generating the proton-motive force for ATP synthesis. Despite remarkable advances in structural knowledge of this complicated membrane-bound enzyme, its mechanism of catalysis remains controversial. In particular, how ubiquinone reduction is coupled to proton pumping and the pathways and mechanisms of proton translocation are contested. We present a 2.4-Å resolution cryo-EM structure of complex I from mouse heart mitochondria in the closed, active (ready-to-go) resting state, with 2945 water molecules modeled. By analyzing the networks of charged and polar residues and water molecules present, we evaluate candidate pathways for proton transfer through the enzyme, for the chemical protons for ubiquinone reduction, and for the protons transported across the membrane. Last, we compare our data to the predictions of extant mechanistic models, and identify key questions to answer in future work to test them.
呼吸复合物 I 是哺乳动物代谢中的一种关键酶，它通过还原 NADH 来捕获泛醌释放的能量，从而驱动质子穿过线粒体内膜，为 ATP 合成产生质子动力。尽管在这个复杂的膜结合酶的结构知识方面取得了显著进展，但它的催化机制仍存在争议。特别是，泛醌还原如何与质子泵浦偶联，以及质子转移的途径和机制都存在争议。我们展示了来自小鼠心脏线粒体的复合物 I 在关闭、活性（准备就绪）静息状态下的 2.4-Å 分辨率冷冻电镜结构，其中建模了 2945 个水分子。通过分析存在的带电和极性残基和水分子网络，我们评估了质子通过酶转移的候选途径，以及泛醌还原的化学质子和跨膜运输的质子。最后，我们将我们的数据与现有的机制模型的预测进行比较，并确定了未来工作中需要回答的关键问题，以对它们进行测试。
